Please wait a minute...
Journal of Integrative Agriculture  2015, Vol. 14 Issue (7): 1390-1397    DOI: 10.1016/S2095-3119(14)60928-X
Animal Science · Veterinary Science Advanced Online Publication | Current Issue | Archive | Adv Search |
Molecular characterization of two candidate genes associated with coat color in Tibetan sheep (Ovis arise)
 HAN Ji-long, YANG Min, GUO Ting-ting, YUE Yao-jing, LIU Jian-bin, NIU Chun-e, WANG Chao-feng, YANG Bo-hui
1、Lanzhou Institute of Husbandry and Pharmaceutical Sciences, Chinese Academy of Agricultural Sciences, Lanzhou 730050,P.R.China
2、Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193, P.R.China
3、College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, P.R.China
Download:  PDF in ScienceDirect  
Export:  BibTeX | EndNote (RIS)      
摘要  Coat color is a key economic trait in sheep. Some candidate genes associated with animal’s coat color were found. Particularly, v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (KIT) and microphthalmia-associated transcription factor (MITF) play a key role in the modulation of hair pigmentation in mammals. This study investigated those two candidate genes’ mutations and expressions associated with wool color in Tibetan sheep. First, the gene polymorphisms of those two genes were analyzed, and then, relative mRNA expression levels of those two genes in skin tissue with different coat colors were compared. Thirdly, KIT and MITF protein expression levels were detected through Western blot and immunehistochemical. Allele C was predominant allele in the white coat color Tibetan sheep population of the MITF coding region g. 1548 C/T loci. The relative MITF mRNA expression in black coat skin tissue was significantly higher than white (P<0.01). However, no significant differences were detected in the KIT gene’s mRNA expression of these two different coat color skin tissues (P>0.05), while the level of KIT protein expression in skin tissues of white and black coats was also roughly equivalent. Our study observed that, the level of MITF protein expression in black coat skin tissue was significantly higher than that in white coat skin tissue, and positive staining for MITF protein expression was detected mainly in the epidermis and the dermal papilla, bulb, and outer root sheath of hair follicles. We conclude that the black coat of Tibetan sheep is related to high MITF expression in the hair follicles, and MITF may be important for coat color formation of Tibetan sheep.

Abstract  Coat color is a key economic trait in sheep. Some candidate genes associated with animal’s coat color were found. Particularly, v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (KIT) and microphthalmia-associated transcription factor (MITF) play a key role in the modulation of hair pigmentation in mammals. This study investigated those two candidate genes’ mutations and expressions associated with wool color in Tibetan sheep. First, the gene polymorphisms of those two genes were analyzed, and then, relative mRNA expression levels of those two genes in skin tissue with different coat colors were compared. Thirdly, KIT and MITF protein expression levels were detected through Western blot and immunehistochemical. Allele C was predominant allele in the white coat color Tibetan sheep population of the MITF coding region g. 1548 C/T loci. The relative MITF mRNA expression in black coat skin tissue was significantly higher than white (P<0.01). However, no significant differences were detected in the KIT gene’s mRNA expression of these two different coat color skin tissues (P>0.05), while the level of KIT protein expression in skin tissues of white and black coats was also roughly equivalent. Our study observed that, the level of MITF protein expression in black coat skin tissue was significantly higher than that in white coat skin tissue, and positive staining for MITF protein expression was detected mainly in the epidermis and the dermal papilla, bulb, and outer root sheath of hair follicles. We conclude that the black coat of Tibetan sheep is related to high MITF expression in the hair follicles, and MITF may be important for coat color formation of Tibetan sheep.
Keywords:  Tibetan sheep       KIT       MITF       gene polymorphism       gene expression       coat color  
Received: 30 May 2014   Accepted:
Fund: 

This work was sponsored by the Earmarked Fund for Modern China Wool & Cashmere Technology Research System, China (CARS-40-03).

Corresponding Authors:  YANG Bo-hui, Tel: +86-931-2115272,E-mail: yangbh2004@163.com     E-mail:  yangbh2004@163.com
About author:  HAN Ji-long, E-mail: hanjilong10000@126.com;

Cite this article: 

HAN Ji-long, YANG Min, GUO Ting-ting, YUE Yao-jing, LIU Jian-bin, NIU Chun-e, WANG Chao-feng, YANG Bo-hui. 2015. Molecular characterization of two candidate genes associated with coat color in Tibetan sheep (Ovis arise). Journal of Integrative Agriculture, 14(7): 1390-1397.

Bai D, Yang L, Unerhu, Zhao Y, Zhao Q, Gaowa H, Manglai D.2011. Effects of KIT gene on coat depigmentation in whitehorses. Hereditas (Beijing), 33, 1171-1178

Bertolotto C, Abbe P, Hemesath T J, Bille K, Fisher D E,Ortonne J P, Ballotti R 1998. Microphthalmia gene productas a signal transducer in cAMP-induced differentiation ofmelanocytes. Journal of Cell Biology, 142, 827-835

Dong C S, Wang H D, Xue L L, Dong Y J, Yang L, Fan R W,Yu X J, Tian X, Ma S H, Smith G W. 2012. Coat colordetermination by miR-137 mediated down-regulation ofmicrophthalmia-associated transcription factor in a mousemodel. RNA, 18, 1679-1686

Fleet M R. 2002. Pigmentation prevention in sheep: Complexor simple? Wool Technology and Sheep Breeding, 50,410-416

Fontanesi L, Scotti E, Russo V. 2012. Haplotype variability inthe bovine MITF gene and association with piebaldism inHolstein and Simmental cattle breeds. Animal Genetics,43, 250-256

Gundry C N, Dobrowolski S F, Martin Y R, Robbins T C, NayL M, Boyd N, Coyne T, Wall M D, Wittwer C T, Teng D.2008. Base-pair neutral homozygotes can be discriminatedby calibrated high-resolution melting of small amplicons.Nucleic Acids Research, 36, 3401-3408

Haase B, Brooks S A, Tozaki T, Burger D, Poncet P A, RiederS, Hasegawa T, Penedo C, Leeb T. 2009. Seven novelKIT mutations in horses with white coat colour phenotypes.Animal Genetics, 40, 623-629

Hauswirth R, Haase B, Blatter M, Brooks S A, Burger D,Drogemuller C, Gerber V, Henke D, Janda J, Jude R. 2012.Mutations in MITF and PAX3 cause “splashed white” andother white spotting phenotypes in horses. Plos Genetics,8, 404-412

Hayes B J, Pryce J, Chamberlain A J, Bowman P J, Goddard ME. 2010. Genetic architecture of complex traits and accuracyof genomic prediction: coat colour, milk-fat percentage,and type in Holstein cattle as contrasting model traits. PlosGenetics, 6, e1001139.

Hemesath T J, Steingrimsson E, McGill G, Hansen M J, VaughtJ, Hodgkinson C A, Arnheiter H, Copeland N G, JenkinsN A, Fisher D E. 1994. Microphthalmia, a critical factor inmelanocyte development, defines a discrete transcriptionfactor family. Genes and Development, 8, 2770-2780

Horikawa T, Norris D A, Johnson T W, Zekman T, DunscombN, Bennion S D, Jackson R L, Morelli J G. 1996. DOPAnegativemelanocytes in the outer root sheath of humanhair follicles express premelanosomal antigens but nota melanosomal antigen or the melanosome-associatedglycoproteins tyrosinase, TRP-1, and TRP-2. Journal ofInvestigative Dermatology, 106, 28-35

Kadekaro A L, Kavanagh R J, Wakamatsu K, Ito S, PipitoneM A, Abdel-Malek Z A. 2003. Cutaneous photobiology.The melanocyte vs. the sun: Who will win the final round?Pigment Cell Research, 16, 434-447

Kijas J W, Lenstra J A, Hayes B, Boitard S, Neto L, SanCristobal M, Servin B, McCulloch R, Whan V, Gietzen K.2012. Genome-wide analysis of the world’s sheep breedsreveals high levels of historic mixture and strong recentselection. Plos Biology, 10, e1001258.

Lalwani A K, Attaie A, Randolph T, Deshmukh D, Wang C,Mhatre A, Wilcox E. 1998. Point mutation in the MITF gene causing Waardenburg syndrome type II in a threegenerationIndian family. American Journal of MedicalGenetics, 80, 406-409

Lee H D, Lee W H, Roh E, Seo C S, Son J K, Lee S H, Hwang BY, Jung S H, Han S B, Kim Y. 2011. Manassantin A inhibitscAMP-induced melanin production by down-regulating thegene expressions of MITF and tyrosinase in melanocytes.Experimental Dermatology, 20, 761-763

Li J Y, Song J S, Bell R, Tran T, Haq R, Liu H F, Love K T, LangerR, Anderson D G, Larue L, Fisher D E. 2012. YY1 regulatesmelanocyte development and function by cooperating withMITF. PLoS Genetics, 8, e1002688.

Limat A, Salomon D, Carraux P, Saurat J H, Hunziker T.1999. Human melanocytes grown in epidermal equivalentstransfer their melanin to follicular outer root sheathkeratinocytes. Archives of Dermatological Research, 291,325-332

Lin J Y, Fisher D E. 2007. Melanocyte biology and skinpigmentation. Nature, 445, 843-850

Lister J A, Robertson C P, Lepage T, Johnson S L, Raible D W.1999. Nacre encodes a zebrafish microphthalmia-relatedprotein that regulates neural crest-derived pigment cell fate.Development, 126, 3757-3767

Liu L, Harris B, Keehan M, Zhang Y. 2009. Genome scan forthe degree of white spotting in dairy cattle. Animal Genetics,40, 975-977

MacKenzie M, Jordan S A, Budd P S, Jackson I J. 1997.Activation of the receptor tyrosine kinase Kit is requiredfor the proliferation of melanoblasts in the mouse embryo.Developmental Biology, 192, 99-107

Nakayama A, Nguyen M, Chen C C, Opdecamp K, HodgkinsonC A, Amheiter H. 1998. Mutations in microphthalmia, themouse homolog of the human deafness gene MITF, affectneuroepithelial and neural crest-derived melanocytesdifferently. Mechanisms of Development, 70, 155-166

Opdecamp K, Vanvooren P, Riviere M, Arnheiter H, MottaR, Szpirer J, Szpirer C. 1998. The rat microphthalmiaassociatedtranscription factor gene (Mitf) maps at 4q34-q41and is mutated in the mib rats. Mammalian Genome, 9,617-621

Otreba M, Rok J, Buszman E, Wrzesniok D. 2012. Regulationof melanogenesis: the role of cAMP and MITF. PostepyHigieny I Medycyny Doswladczalnej, 66, 33-40

 (in Polish)Penagaricano F, Zorrilla P, Naya H, Robello C, Urioste J I. 2012.Gene expression analysis identifies new candidate genesassociated with the development of black skin spots inCorriedale sheep. Journal of Applied Genetics, 53, 99-106

Peters E, Tobin D J, Botchkareva N, Maurer M, Paus R. 2002.Migration of melanoblasts into the developing murine hairfollicle is accompanied by transient c-Kit expression. Journalof Histochemistry & Cytochemistry, 50, 751-766

Philipp U, Lupp B, Momke S, Stein V, Tipold A, Eule J C,Rehage J, Distl O. 2011. A MITF mutation associated with adominant white phenotype and bilateral deafness in GermanFleckvieh cattle. PLoS One, 6, e28857.

Royo L J, Alvarez I, Arranz J J, Fernandez I, Rodriguez A,Perez-Pardal L, Goyache F. 2008. Differences in theexpression of the ASIP gene are involved in the recessiveblack coat colour pattern in sheep: evidence from the rareXalda sheep breed. Animal Genetics, 39, 290-293

Shoag J, Haq R, Zhang M, Liu L, Rowe G C, Jiang A, KoulisisN, Farrel C, Amos C I, Wei Q. 2013. PGC-1 coactivatorsregulate MITF and the tanning response. Molecular Cell,49, 145-157

Steingrimsson E, Copeland N G, Jenkins N A. 2004.Melanocytes and the microphthalmia transcription factornetwork. Annual Review of Genetics, 38, 365-411

Takeda K, Yasumoto K, Kawaguchi N, Udono T, WatanabeK, Saito H, Takahashi K, Noda M, Shibahara S. 2002.Mitf-D, a newly identified isoform, expressed in the retinalpigment epithelium and monocyte-lineage cells affectedby Mitf mutations. Biochimica et Biophysica Acta-GeneStructure and Expression, 1574, 15-23

Taylor B A, Navin A, Phillips S J. 1994. PCR-Amplification ofsimple sequence repeat variants from pooled DNA samplesfor rapidly mapping new mutations of the mouse. Genomics,21, 626-632

Vachtenheim J, Borovansky J. 2010. “Transcription physiology”of pigment formation in melanocytes: central role of MITF.Experimental Dermatology, 19, 617-627

Yasumoto K, Amae S, Udono T, Fuse N, Takeda K, ShibaharaS. 1998. A big gene linked to small eyes encodes multipleMitf isoforms: Many promoters make light work. PigmentCell Research, 11, 329-336

Zhang R, Fan R, Cheng Z, Tian X, Liu J, Gao L, Ma Z, Dong C.2011. Regulation of TYR and MITF mRNA expression byCDK5 in alpaca melanocytes. Chinese Journal of Animaland Veterinary Sciences, 42, 1712-1717 (in Chinese)

Zhu Z, He J, Jia X, Jiang J, Bai R, Yu X, Lv L, Fan R, He X,Geng J. 2010. MicroRNA-25 functions in regulation ofpigmentation by targeting the transcription factor MITF inalpaca (Lama pacos) skin melanocytes. Domestic AnimalEndocrinology, 38, 200-209
[1] ZHAO Shu-ping, DENG Kang-ming, ZHU Ya-mei, JIANG Tao, WU Peng, FENG Kai, LI Liang-jun.

Optimization of slow-release fertilizer application improves lotus rhizome quality by affecting the physicochemical properties of starch [J]. >Journal of Integrative Agriculture, 2023, 22(4): 1045-1057.

[2] ZHANG Yan-mei, AO De, LEI Kai-wen, XI Lin, Jerry W SPEARS, SHI Hai-tao, HUANG Yan-ling, YANG Fa-long. Dietary copper supplementation modulates performance and lipid metabolism in meat goat kids[J]. >Journal of Integrative Agriculture, 2023, 22(1): 214-221.
[3] JIANG Yong, MA Xin-yan, XIE Ming, ZHOU Zheng-kui, TANG Jing, CHANG Guo-bin, CHEN Guo-hong, HOU Shui-sheng. Dietary threonine deficiency affects expression of genes involved in lipid metabolism in adipose tissues of Pekin ducks in a genotype-dependent manner[J]. >Journal of Integrative Agriculture, 2022, 21(9): 2691-2699.
[4] RONG Hao, YANG Wen-jing, XIE Tao, WANG Yue, WANG Xia-qin, JIANG Jin-jin, WANG You-ping. Transcriptional profiling between yellow- and black-seeded Brassica napus reveals molecular modulations on flavonoid and fatty acid content[J]. >Journal of Integrative Agriculture, 2022, 21(8): 2211-2226.
[5] AN Feng, ZHANG Kang, ZHANG Ling-kui, LI Xing, CHEN Shu-min, WANG Hua-sen, CHENG Feng. Genome-wide identification, evolutionary selection, and genetic variation of DNA methylation-related genes in Brassica rapa and Brassica oleracea[J]. >Journal of Integrative Agriculture, 2022, 21(6): 1620-1632.
[6] FAN Xiao-xue, BIAN Zhong-hua, SONG Bo, XU Hai. Transcriptome analysis reveals the differential regulatory effects of red and blue light on nitrate metabolism in pakchoi (Brassica campestris L.)[J]. >Journal of Integrative Agriculture, 2022, 21(4): 1015-1027.
[7] LIU Cong, LI De-xiong, HUANG Xian-biao, Zhang Fu-qiong, Xie Zong-zhou, Zhang Hong-yan, Liu Ji-hong. Manual thinning increases fruit size and sugar content of Citrus reticulata Blanco and affects hormone synthesis and sugar transporter activity[J]. >Journal of Integrative Agriculture, 2022, 21(3): 725-735.
[8] DUAN Yao-ke, HAN Rong, SU Yan, WANG Ai-ying, LI Shuang, SUN Hao, GONG Hai-jun. Transcriptional search to identify and assess reference genes for expression analysis in Solanum lycopersicum under stress and hormone treatment conditions[J]. >Journal of Integrative Agriculture, 2022, 21(11): 3216-3229.
[9] Kashif NOOR, Hafiza Masooma Naseer CHEEMA, Asif Ali KHAN, Rao Sohail Ahmad KHAN. Expression profiling of transgenes (Cry1Ac and Cry2A) in cotton genotypes under different genetic backgrounds[J]. >Journal of Integrative Agriculture, 2022, 21(10): 2818-2832.
[10] WANG Pei-pei, WANG Zhao-ke, GUAN Le, Muhammad Salman HAIDER, Maazullah NASIM, YUAN Yong-bing, LIU Geng-sen, LENG Xiang-peng. Versatile physiological functions of the Nudix hydrolase family in berry development and stress response in grapevine[J]. >Journal of Integrative Agriculture, 2022, 21(1): 91-112.
[11] GUO Bing-bing, LI Jia-ming, LIU Xing, QIAO Xin, Musana Rwalinda FABRICE, WANG Peng, ZHANG Shao-ling, WU Ju-you. Identification and expression analysis of the PbrMLO gene family in pear, and functional verification of PbrMLO23[J]. >Journal of Integrative Agriculture, 2021, 20(9): 2410-2423.
[12] JI Xiao-hao, WANG Bao-liang, WANG Xiao-di, WANG Xiao-long, LIU Feng-zhi, WANG Hai-bo. Differences of aroma development and metabolic pathway gene expression between Kyoho and 87-1 grapes[J]. >Journal of Integrative Agriculture, 2021, 20(6): 1525-1539.
[13] CHEN Chang-long, YUAN Fang, LI Xiao-ying, MA Rong-cai, XIE Hua. Jasmonic acid and ethylene signaling pathways participate in the defense response of Chinese cabbage to Pectobacterium carotovorum infection[J]. >Journal of Integrative Agriculture, 2021, 20(5): 1314-1326.
[14] PENG Chao, MA Biao, ZHANG Chen. Poverty alleviation through e-commerce: Village involvement and demonstration policies in rural China[J]. >Journal of Integrative Agriculture, 2021, 20(4): 998-1011.
[15] WANG Lu-lu, ZHAO Chun-fang, LIU Chang-jun, ZHANG Hao, LIAN Ling. Analysis of DNA methylation of CD79B in MDV-infected chicken spleen[J]. >Journal of Integrative Agriculture, 2021, 20(11): 2995-3002.
No Suggested Reading articles found!